Syringe pump system for pulse-free metering and precise mixing in hplc uhplc, micro-hplc and nano-hplc

ABSTRACT

The invention describes a high-pressure syringe pump system for use in the field of HPLC, UHPLC, micro-HPLC and nano-HPLC in gradient operation. The different solvents are compressed independently by passive non-return valves, the solvent combination does not take place until at an identical pressure, and the target pressure for the initialization is achieved by a control setting with a higher delivery capacity, in which the compressibility is also taken into consideration. The changing of solvents and the flushing of the pump heads take place by way of separate inlets and outlets in the throughflow via an active multiple-position high-pressure valve which can also close the pump head and can open it via the passive non-return valve. The regulating unit and the drive motor which also serves as force sensor detect compressibility, air inclusions and leaks. Depending on the embodiment, the syringe pump heads are stabilized thermally.

The invention relates to a syringe pump system in the field of HPLC, UHPLC, as well as nano-HPLC and micro-HPLC, for pulse-free conveying and precise mixing of solvents, and to a method of quickly adjusting mixing ratios of different solvents having different compressibilities, even at very high pressure, to improve the repeat accuracy of gradients and improve handling, and to allow new uses.

In HPLC there is a trend toward ever greater sample throughput at ever smaller sample volumes. Accordingly, “Ultra High Performance Liquid Chromatography” (UHPLC) is understood to be HPLC with greatly increased performance. Reduction of the column diameter goes even further in its development and has led, beyond that, to the development of nano-HPLC and micro-HPLC.

For conveying solvents in the field of HPLC, UHPLC, as well as nano-HPLC and micro-HPLC, great efforts have been made for a long time to achieve precise and reproducible results with the least possible effort.

Conventional pump systems for the field of HPLC, UHPLC, as well as nano-HPLC and micro-HPLC, are very complicated and are generally built for continuous throughput. They therefore require special provisions for damping the pulsation that is caused by the pump piston movements. Such pulsations are deviations from the average pressure flow and lead to pressure variations in the HPLC system. Particularly in the case of nano-HPLC and micro-HPLC, even small pressure variations can cause significant errors.

Furthermore, the different compressibility of solvents used, which are different from one another, or of solvents having a different gas saturation, has a direct influence on the mixing precision and on the time required for system equilibration, particularly in the case of low flow rates of a few nl/min to 5 ml/min and high pressures of up to more than 1000 bar.

Conventional syringe pumps are generally not constructed for gradient operation, small flow rates, and for high pressures.

The compressibility of the solvents and the mixing precision are therefore not taken into consideration or are only taken into consideration very insufficiently.

In gradient elution with a high-pressure gradient, two pumps are generally used. In the gradient design, the type of solvent selected plays a major role in mixing precision. If, for example, methanol/water mixtures are used, the mixing reaction is already highly exothermal. Volume reduction occurs, along with a viscosity increase and pressure increase.

In a different gradient design, such as acetonitrile and water, for example, such effects are not as strongly present and therefore less serious.

Great demands are therefore made on the pump system. Thus, a constant and reproducible flow velocity should be guaranteed, because the reproducibility of the measurement depends on it. Furthermore, the most pulse-free flow possible must be guaranteed. Pressure surges can actually damage the stationary phase.

In conventional systems, the problems presented cannot be averted and taken into consideration, or can only be averted and taken into consideration insufficiently.

Proceeding from this, the object of invention is to provide a syringe pump system for HPLC that works essentially in a pulse-free manner, is suitable for gradient operation, and carries out a desired gradient profile in precise and reproducible manner, even at very high pressure.

This object is attained by a syringe pump system for HPLC having at least two solvents, in which two syringe pumps, independent of one another, are coupled to form a binary system, wherein each syringe pump has a pump cylinder that is connected with a respective multi-position valve through a respective outlet connection line that is provided with a respective passive pump outlet valve, a respective intake line, and a respective washing outlet line, wherein each multi-position valve has a further outlet line, in each instance that ends in a single outlet line by manifold fitting, and the pressure required for chromatographic analysis, for each of the solvents, is built up separately in the respective pump cylinder, by the pump piston, on the basis of the passive pump outlet valve.

According to the invention, the different solvents required for gradient operation are therefore compressed independently by the respective passive pump outlet valves as passive check valves, so that combining the solvents by the manifold fitting, which can be a T-fitting, for example, only takes place when they are at the same pressure.

In the syringe pump system according to the invention, the target pressure is quickly achieved by a default with a higher pump output.

Furthermore, it can be provided in the syringe pump system that each of the syringe pumps has a motor as a drive for the piston and as a power sensor for the compression of the respective solvent.

Preferably, a controller is additionally provided, and the motor together with the controller detects air inclusions, the compressibility of the solvent, and leaks.

The invention also relates to a method of pulse-free conveying and precise mixing of at least two solvents for HPLC in gradient operation, in which method, in a syringe pump system having two syringe pumps independent of one another, each of the pump cylinders is connected with a multi-position valve, is flushed and closed by this valve by lines separated from one another, and is operated by a passive pump outlet valve, so that the pressure buildup required for gradient operation takes place in the respective pump cylinder, and mixing of the solvents only takes place when the same pressure has been reached, in a manifold fitting.

In this connection, changing of the solvent as well as flushing of the syringe pump cylinders takes place by separate inlets and outlets in the through-flow, by the multi-position high-pressure valve that can close off the pump cylinder and open it by the passive pump outlet valve or check valve. For this reason, the multi-position high-pressure valve is also referred to as an active multi-position high-pressure valve.

Preferably, the pressure builds up synchronously in the respective syringe pumps.

Furthermore, the pistons of the syringe pumps are driven by respective motors whose motor force is measured separately, and the solvent pressure in the pump cylinders is essentially proportional to the motor force.

In this connection, the drive motor can serve as a force sensor and can also have a high-resolution position detection unit and position controller.

Preferably, the syringe pumps are re-initialized for each individual HPLC analysis, so that as a result, the same starting conditions exist for the syringe pumps and the chromatographic analysis.

The invention furthermore relates to the use of the syringe pump system as described above, and of the method in its different embodiments in the field of HPLC, UHPLC, as well as nano-HPLC and micro-HPLC.

In this connection, investigations with gradients and essentially pulse-free flow for HPLC columns at 2.1 mm to 150 μm are possible.

However, the syringe pump system according to the invention, in its embodiments, can be used not only for gradient elution, but rather can be used just as well for isocratic elution with two isocratic pumps that are independent of one another, and in continuous flow mode.

In the following, the invention will be explained in greater detail using an illustrated embodiment, making reference to FIG. 1 of the drawing.

The figures show:

FIG. 1 is a schematic view of a binary syringe pump system according to the invention,

FIG. 2 a is a detail like FIG. 1 that shows an active multi-position valve in a position for an ejection movement of the syringe piston during the flushing procedure of the pump cylinder,

FIG. 2 b is a detail like FIG. 1 that shows an active multi-position valve in a position for an intake movement of the syringe piston during the flushing procedure of the pump cylinder,

FIG. 2 c is a detail like FIG. 1 that shows an active multi-position valve in a position in which the pump cylinder is closed, and

FIG. 2 d is a detail like FIG. 1 that shows an active multi-position valve in a position in which a solvent can be pumped into an outlet connection line and via a mixing T-fitting, through an outlet line.

In FIG. 1, a binary syringe pump system having syringe pumps is shown that is suitable for gradient operation and has two pumps 1 and 1′ that are separate from and independent of one another. These two pumps 1 and 1′ in turn each have drives, referred to hereinafter as motors 3 and 3′, having respective ball screws 5 and 5′ that forms the respective motor axles and axially moves respective ball nuts 7 and 7′ of respective drive carriages 9 and 9′ and pump pistons 11 and 11′.

Pump cylinders 13 and 13′ are sealed by respective seals 15 and 15′. The pump cylinders 13 and 13′ are connected with respective valves by respective intake lines 17 and 17′, respective washing outlet lines 19 and 19′, respective passive pump outlet valves 21 and 21′, and respective outlet connection lines 23 and 23′, which valves are here referred to as active multi-position valves 25 and 25′.

The active multi-position valves 25 and 25′ have respective rotors and four possible positions of respective connection grooves 27 and 27′ shown in FIGS. 2 a-2 d and explained below. In this connection, reference is first made to FIGS. 2 a and 2 b that describe a flushing procedure for the pump cylinders 13 and 13′. The flushing procedure takes place to start with by an ejection movement of the pump pistons 11 and 11′, in that the multi-position valves 25 shifts the connection groove 27 to position A in FIG. 2 a, and thereafter by an intake movement of the pump pistons 11 and 11′ in which the connection groove 27 of the multi-position valve 25 is shifted to position B in FIG. 2 b. For this purpose, flushing intake connections 29 and 29′ and flushing ejection connections 31 and 31′ are provided on the respective pump cylinders 13 and 13′, at the opposite ends thereof. As a result, flow in one direction takes place in them during the flushing procedure.

In FIG. 2 c, a further position is shown in which the connection grooves 27 or 27′ of the multi-position valves 25 and 25′ are in position C in FIG. 2 c. In this position, the pump cylinders 20 and 20′ [13 and 13′] are closed, and now a possible air inclusion, the compressibility of the solvent, as well as the tightness of the piston seals 15 and 15′, of the flushing intake connections 29 and 29′, and of the flushing ejection connections 31 and 31′, of the intake lines 17 and 17′, of the outlet connection line 23 and 23′, of the washing outlet lines 19 and 19′, of the connections at the passive pump outlet valves 21 and 21′, and of the multi-position valves 25 and 25′ are detected by the motor controller and the force measurement of the motors 3 and 3′.

In this manner, the motors 3 and 3′ simultaneously serve as force sensors that, together with a controller, detect the compressibility and possible leaks during closure of the respective pump cylinders 20 or 20′ in the position C of the connection grooves 27 or 27′ of the multi-position valves 25 or 25′. The controller is configured as a high-resolution position detection unit and position controller that is not shown again separately as such.

Only when the connection grooves 27 and 27′ of the multi-position valves 25 and 25′ are in position D in FIG. 2 d can the solvent be pressed into a mixing T-fitting 35 and through a common outlet line 37, with a positive piston stroke, by the passive pump outlet valves 21 and 21′ of the respective outlet connection lines 23 and 23′ and the respective further outlet lines 33 and 33′.

In this connection, depending on the chromatographic HPLC method being used, pressures of up to more than 1000 bar in the outlet line 37 must be expected. The passive pump outlet valves 21 and 21′ ensure that at different compressibility in the respective pump cylinders 13 and 13′ of the respective solvents used, the solvents are compressed only by the respective pump pistons 11 and 11′, and mixing in the mixing T-fitting 35 of the two solvents only takes place when the same pressure is reached. The time for reaching the chromatographic starting conditions is dependent on the pressure, the mixing ratio, the flow velocity, as well as the compressibility of the different solvents. For this reason, the pressure will build up synchronously in the pump cylinders 13 and 13′.

According to the invention, the chromatographic starting conditions can be reached significantly more quickly and reliably than is conventionally possible in the state of the art. This is basically due to the fact that within the scope of the present invention, separate motor-force measurement of the drive motors 3 and 3′ is carried out as explained above. In addition, the solvent pressure in the pump cylinders 13 and 13′ is extensively proportional to the motor force in the syringe pump system according to the invention. In this manner, the chromatographic starting conditions are reached significantly more quickly by corresponding motor regulation and a starting pressure default.

The different solvents required for gradient operation in the binary syringe pump system according to the invention are compressed independent of one another in the two pumps 1 and 1′ that are separate from and independent of each other, using the passive pump outlet valves 21, 21′ that act as check valves, so that combining the solvents only takes place when they are at the same pressure. In this connection, the desired target pressure is quickly reached by a default with a higher pump output.

Because of the fact that the pump cylinders 13 and 13′ are connected with the respective active multi-position valve 25 and 25′ by the respective intake lines 17 and 17′, the respective washing outlet lines 19 and 19′, the respective passive pump outlet valves 21 and 21′, and the respective outlet connection lines 23 and 23′, changing solvents as well as flushing the pump cylinders 13 and 13′ can take place by separate inlets and outlets in the through-flow, by the multi-position valve 25 and 25′ that can also be referred to as a multi-position high-pressure valve 25 and 25′. They are referred to as active because, among other things, they can close the respective pump cylinders 13 and 13′ connected to them and can open them by the respective passive check valve.

On the basis of the structure of the binary syringe pump system according to the invention, as explained, in which each individual pump cylinder 13 and 13′ is flushed and closed by the active multi-position valve 25 and 25′, by the separate lines, and operated by the passive pump outlet valve 21 and 21′ that is configured as a check valve, and furthermore, the flushing intake connection and the washing outlet line 19 and 19′ as an outlet flushing connection are situated on the respective pump cylinder 13 and 13′, separately from one another, and, in total, pressure buildup can take place in the respective pump cylinder 13 and 13′, only using the pump piston 11 and 11′, the result is likewise achieved that the system works in essentially pulse-free manner.

In addition, depending on the desired embodiment, the pump cylinders 13 and 13′ of the syringe pump system can be thermally stabilized. Materials that do not change and thereby distort the desired analytical results are used for the pump cylinders 13 and 13′.

Particularly in the case of low pump output, it is advantageous if the pump cylinders 13 and 13′ are thermally stabilized and if the pressure lines and the multi-position valve are insulated.

It is furthermore possible that multiple pump outlets are connected together so that reaching the chromatographic starting conditions of each analysis or each analysis run takes place by a regulation characteristic in which the force sensors of each of the syringe pumps takes into account the pressure buildup and the conveying velocity of all the coupled pumps. 

1. A syringe pump system for HPLC with at least two solvents, having two mutually independent syringe pumps coupled to form a binary system and each having a pump cylinder that is connected with a multi-position valve by an outlet connection line that is provided with a respective passive pump outlet valve, a respective intake line, and a respective washing outlet line, each multi position valve having a further outlet line that opens into a common outlet line at a manifold fitting, the pressure required for chromatographic analysis, for each of the solvents, being built up separately in the respective pump cylinders, by the respective pump pistons and on the basis of the respective passive pump outlet valve.
 2. The syringe pump system according to claim 1, wherein each syringe pump has a motor as a drive for the respective pump piston and as a force sensor for compression of the respective solvent.
 3. The syringe pump system according to claim 2, wherein in addition a controller is provided, and that the motor and the controller detect air inclusions, the compressibility of the solvent, and leaks.
 4. A method of pulse-free feeding and precise mixing of at least two solvents for HPLC in gradient operation, using a syringe pump system having two mutually independent syringe pumps each having respective pump cylinders connected with respective multi-position valves, flushed and closed by this valve by lines separate from one another, and operated by a passive pump outlet valve so that the pressure buildup required for gradient operation takes place in the respective pump cylinders and mixing of the solvents in a manifold only takes place when the same pressure has been reached.
 5. The method according to claim 4, wherein the pressure in the syringe pumps builds up synchronously.
 6. The method according to claim 4, wherein the pistons of the syringe pumps are driven by respective motors whose motor force is measured separately, and that the solvent pressure in the pump cylinders is essentially proportional to the motor force.
 7. The method according to claim 4, wherein the syringe pumps are re-initialized for each individual HPLC analysis, and thereby the same starting conditions exist for the syringe pumps and chromatographic analysis.
 8. Use of the syringe pump system according to claim 1 in the field of HPLC, UHPLC, as well as nano-HPLC and micro-HPLC.
 9. Use of the syringe pump system according to claim 1 for gradient elution, isocratic elution, and in continuous flow mode. 